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Structural organization of the dynein-dynactin complex bound to microtubules.

Chowdhury S, Ketcham SA, Schroer TA, Lander GC - Nat. Struct. Mol. Biol. (2015)

Bottom Line: Cytoplasmic dynein associates with dynactin to drive cargo movement on microtubules, but the structure of the dynein-dynactin complex is unknown.Using electron microscopy, we determined the organization of native bovine dynein, dynactin and the dynein-dynactin-microtubule quaternary complex.In the microtubule-bound complex, the dynein motor domains are positioned for processive unidirectional movement, and the cargo-binding domains of both dynein and dynactin are accessible.

View Article: PubMed Central - PubMed

Affiliation: Department of Integrative Structural and Computational Biology, Scripps Research Institute, La Jolla, California, USA.

ABSTRACT
Cytoplasmic dynein associates with dynactin to drive cargo movement on microtubules, but the structure of the dynein-dynactin complex is unknown. Using electron microscopy, we determined the organization of native bovine dynein, dynactin and the dynein-dynactin-microtubule quaternary complex. In the microtubule-bound complex, the dynein motor domains are positioned for processive unidirectional movement, and the cargo-binding domains of both dynein and dynactin are accessible.

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Organization of the DDB-MT complex(A) Two class averages of MT-bound DDB complexes, attached at slightly different angles relative to the MT. (B) 2D averages in the same orientation as (A) overlaid with a low-pass filtered 3D dynactin model (from Fig. 2) colored blue. (C) Dynein components in the 2D averages are assigned based on information determined from the isolated dynein class averages (Fig. 1). HCs are yellow, LICs are orange, the IC C-term WD40 domains are blue, and the putative LC7 dimer is green. (D) Two class averages of DDB-MT complexes showing the location of the shoulder and an extension (labeled “p150 arm”) that wraps around the Arp1 filament and contacts dynein. We propose this corresponds to the p150Glued coiled-coil. (E) A model of the DDB complex in the same orientation as the averages in (D), with the path of the putative p150Glued coiled-coil extension (“p150 arm”) traced in light blue. Scale bar in (A) corresponds to 25 nm, and all EM images are at the same scale.
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Figure 3: Organization of the DDB-MT complex(A) Two class averages of MT-bound DDB complexes, attached at slightly different angles relative to the MT. (B) 2D averages in the same orientation as (A) overlaid with a low-pass filtered 3D dynactin model (from Fig. 2) colored blue. (C) Dynein components in the 2D averages are assigned based on information determined from the isolated dynein class averages (Fig. 1). HCs are yellow, LICs are orange, the IC C-term WD40 domains are blue, and the putative LC7 dimer is green. (D) Two class averages of DDB-MT complexes showing the location of the shoulder and an extension (labeled “p150 arm”) that wraps around the Arp1 filament and contacts dynein. We propose this corresponds to the p150Glued coiled-coil. (E) A model of the DDB complex in the same orientation as the averages in (D), with the path of the putative p150Glued coiled-coil extension (“p150 arm”) traced in light blue. Scale bar in (A) corresponds to 25 nm, and all EM images are at the same scale.

Mentions: In the DDB-MT complex, dynein is aligned between dynactin and the MT with the distal part of the dynein tail near the dynactin pointed end. The dynein tail associates with the short strand of the dynactin Arp filament, on the side opposite the shoulder (Fig. 3), consistent with the recently determined structure of the isolated dynein tail-dynactin-BicD2N complex20. Densities that cannot be attributed to either dynactin or dynein likely correspond to the BicD2N coiled-coil. Notably, the centers of the dynein heads are approximately 17 nm away from the MT and the MT-binding stalks are oriented at an acute angle (Supplementary Fig. 5d,e). Unlike the motor domains in free DDB complexes12,21, which exhibit a range of orientations and separation, those in DDB-MT complexes are in close proximity to one another (about 7 nm apart) but are not locked into a single orientation relative to each other (Supplementary Fig. 5e,g). The adjacent HC kink may provide a flexible “shock absorber” that allows the motor domains to undergo the structural changes that underlie stepping without interfering with dynactin and cargo interactions. The organization of dynein and dynactin in the DDB-MT complex also elucidates the arrangement of cargo-binding domains. Using the isolated dynein class averages for comparison, densities corresponding to the IC-LC dimerization domain and the LICs (Fig. 3b, green and orange, respectively) are apparently exposed in the DDB-MT complex. Given the importance of these regions for interactions with dynactin and other binding partners, it makes sense that these parts of the tail remain accessible.


Structural organization of the dynein-dynactin complex bound to microtubules.

Chowdhury S, Ketcham SA, Schroer TA, Lander GC - Nat. Struct. Mol. Biol. (2015)

Organization of the DDB-MT complex(A) Two class averages of MT-bound DDB complexes, attached at slightly different angles relative to the MT. (B) 2D averages in the same orientation as (A) overlaid with a low-pass filtered 3D dynactin model (from Fig. 2) colored blue. (C) Dynein components in the 2D averages are assigned based on information determined from the isolated dynein class averages (Fig. 1). HCs are yellow, LICs are orange, the IC C-term WD40 domains are blue, and the putative LC7 dimer is green. (D) Two class averages of DDB-MT complexes showing the location of the shoulder and an extension (labeled “p150 arm”) that wraps around the Arp1 filament and contacts dynein. We propose this corresponds to the p150Glued coiled-coil. (E) A model of the DDB complex in the same orientation as the averages in (D), with the path of the putative p150Glued coiled-coil extension (“p150 arm”) traced in light blue. Scale bar in (A) corresponds to 25 nm, and all EM images are at the same scale.
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Related In: Results  -  Collection

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Figure 3: Organization of the DDB-MT complex(A) Two class averages of MT-bound DDB complexes, attached at slightly different angles relative to the MT. (B) 2D averages in the same orientation as (A) overlaid with a low-pass filtered 3D dynactin model (from Fig. 2) colored blue. (C) Dynein components in the 2D averages are assigned based on information determined from the isolated dynein class averages (Fig. 1). HCs are yellow, LICs are orange, the IC C-term WD40 domains are blue, and the putative LC7 dimer is green. (D) Two class averages of DDB-MT complexes showing the location of the shoulder and an extension (labeled “p150 arm”) that wraps around the Arp1 filament and contacts dynein. We propose this corresponds to the p150Glued coiled-coil. (E) A model of the DDB complex in the same orientation as the averages in (D), with the path of the putative p150Glued coiled-coil extension (“p150 arm”) traced in light blue. Scale bar in (A) corresponds to 25 nm, and all EM images are at the same scale.
Mentions: In the DDB-MT complex, dynein is aligned between dynactin and the MT with the distal part of the dynein tail near the dynactin pointed end. The dynein tail associates with the short strand of the dynactin Arp filament, on the side opposite the shoulder (Fig. 3), consistent with the recently determined structure of the isolated dynein tail-dynactin-BicD2N complex20. Densities that cannot be attributed to either dynactin or dynein likely correspond to the BicD2N coiled-coil. Notably, the centers of the dynein heads are approximately 17 nm away from the MT and the MT-binding stalks are oriented at an acute angle (Supplementary Fig. 5d,e). Unlike the motor domains in free DDB complexes12,21, which exhibit a range of orientations and separation, those in DDB-MT complexes are in close proximity to one another (about 7 nm apart) but are not locked into a single orientation relative to each other (Supplementary Fig. 5e,g). The adjacent HC kink may provide a flexible “shock absorber” that allows the motor domains to undergo the structural changes that underlie stepping without interfering with dynactin and cargo interactions. The organization of dynein and dynactin in the DDB-MT complex also elucidates the arrangement of cargo-binding domains. Using the isolated dynein class averages for comparison, densities corresponding to the IC-LC dimerization domain and the LICs (Fig. 3b, green and orange, respectively) are apparently exposed in the DDB-MT complex. Given the importance of these regions for interactions with dynactin and other binding partners, it makes sense that these parts of the tail remain accessible.

Bottom Line: Cytoplasmic dynein associates with dynactin to drive cargo movement on microtubules, but the structure of the dynein-dynactin complex is unknown.Using electron microscopy, we determined the organization of native bovine dynein, dynactin and the dynein-dynactin-microtubule quaternary complex.In the microtubule-bound complex, the dynein motor domains are positioned for processive unidirectional movement, and the cargo-binding domains of both dynein and dynactin are accessible.

View Article: PubMed Central - PubMed

Affiliation: Department of Integrative Structural and Computational Biology, Scripps Research Institute, La Jolla, California, USA.

ABSTRACT
Cytoplasmic dynein associates with dynactin to drive cargo movement on microtubules, but the structure of the dynein-dynactin complex is unknown. Using electron microscopy, we determined the organization of native bovine dynein, dynactin and the dynein-dynactin-microtubule quaternary complex. In the microtubule-bound complex, the dynein motor domains are positioned for processive unidirectional movement, and the cargo-binding domains of both dynein and dynactin are accessible.

Show MeSH
Related in: MedlinePlus